Virtual Anthropology: forensic applications to cranial skeletal remains from the Spanish Civil War

Biological and forensic anthropologists face limitations while studying skeletal remains altered by taphonomic alterations and perimortem trauma, such as in remains from the Spanish Civil War. However, virtual anthropology techniques can optimize the information inferred from fragmented and deformed remains by generating and restoring three-dimensional bone models. We applied a low-cost 3D modelling methodology based on photogrammetry to develop novel forensic applications of virtual 3D skull reconstruction, assembly, restoration and ancestry estimation. Crania and mandible fragments from five Spanish Civil War victims were reconstructed with high accuracy, and only one cranium could not be assembled due to extensive bone loss. Virtual mirroring successfully restored reconstructed crania, producing 3D models with reduced deformation and perimortem trauma. High correlation between traditional and virtual craniofacial measurements confirmed that 3D models are suitable for forensic applications. Craniometric databases of world-wide and Spanish populations were used to assess the potential of discriminant analysis to estimate population ancestry. Our protocol correctly estimated the continental origin of 86.7 % of 15 crania of known origin, and despite low morphological differentiation within European populations, correctly identified 54.5 % as Spanish and 27.3 % of them with high posterior probabilities. Two restored crania from the Civil War were estimated as Spanish, and one as a non-Spanish European. Results were not conclusive for one cranium and did not confirm previous archeological hypotheses. Overall, our research shows the potential to assess the presence of foreign volunteers in the Spanish Civil War and highlights the added value of 3D-virtual techniques in forensic anthropology

µMatch: 3D shape correspondence for biological image data

Modern microscopy technologies allow imaging biological objects in 3D over a wide range of spatial and temporal scales, opening the way for a quantitative assessment of morphology. However, establishing a correspondence between objects to be compared, a first necessary step of most shape analysis workflows, remains challenging for soft-tissue objects without striking features allowing them to be landmarked. To address this issue, we introduce the μMatch 3D shape correspondence pipeline. μMatch implements a state-of-the-art correspondence algorithm initially developed for computer graphics and packages it in a streamlined pipeline including tools to carry out all steps from input data pre-processing to classical shape analysis routines. Importantly, μMatch does not require any landmarks on the object surface and establishes correspondence in a fully automated manner. Our open-source method is implemented in Python and can be used to process collections of objects described as triangular meshes. We quantitatively assess the validity of μMatch relying on a well-known benchmark dataset and further demonstrate its reliability by reproducing published results previously obtained through manual landmarking

Phenotyping the brain, the face, and their genetic interaction over development

The development of the brain and the face is intimately coordinated through a continuous physical and molecular interaction during morphogenesis. Understanding how dynamic spatio-temporal regulation of gene expression patterns guide this process is crucial to reveal mechanisms that may have contributed to human evolution

Green Tea catechins modulate skeletal development with effects dependent on dose, time, and structure in a Down syndrome mouse model

Altered skeletal development in Down syndrome (DS) results in a brachycephalic skull, flattened face, shorter mandibular ramus, shorter limbs, and reduced bone mineral density (BMD). Our previous study showed that low doses of green tea extract enriched in epigallocatechin-3-gallate (GTE-EGCG), administered continuously from embryonic day 9 to postnatal day 29, reduced facial dysmorphologies in the Ts65Dn (TS) mouse model of DS, but high doses could exacerbate them. Here, we extended the analyses to other skeletal structures and systematically evaluated the effects of high and low doses of GTE-EGCG treatment over postnatal development in wild-type (WT) and TS mice using in vivo µCT and geometric morphometrics. TS mice developed shorter and wider faces, skulls, and mandibles, together with shorter and narrower humerus and scapula, and reduced BMD dynamically over time. Besides facial morphology, GTE-EGCG did not rescue any other skeletal phenotype in TS treated mice. In WT mice, GTE-EGCG significantly altered the shape of the skull and mandible, reduced the length and width of the long bones, and lowered the BMD. The disparate effects of GTE-EGCG depended on the dose, developmental timepoint, and anatomical structure analyzed, emphasizing the complex nature of DS and the need to further investigate the simultaneous effects of GTE-EGCG supplementation

Facial Biomarkers Detect Gender-Specific Traits for Bipolar Disorder

Bipolar disorder (BD) is a psychiatric disorder associated with brain and neurodevelopmental alterations. As in other disorders, patients with BD present minor Physical Anomalies (MPAs) in higher frequency than healthy subjects. MPAs are subtle signs of developmental deviation that appear in body regions that share the ectodermal origin of the brain and are likely triggered by the same insults altering early brain development in mental disorders. MPAs are thus considered potential biomarkers for neurodevelopmental disorders. In this study, we compared facial shape variation between patients with BD and healthy controls using 3D facial reconstructions from magnetic resonance images (MRI) to test the potential of MPAs as a biomarker of BD diagnosis. Moreover, we assessed sex-specific facial shape variation to test whether the disorder affects differently male and female patients. We collected the 3D coordinates of 20 anatomical facial landmarks in a sample of 174 subjects (87 patients with BD and 87 healthy controls) and analyzed global and local patterns of facial shape using Geometric Morphometrics and multivariate statistical techniques. Although Procrustes-ANOVA analysis revealed that diagnosis accounted for a low but significant effect (1.1% of total facial shape variance, P-value=0.016), global facial shape did not significantly discriminate between patients with BD and healthy controls (P-value=0.19). However, Euclidean Distance Matrix Analysis (EDMA) based on local distances of the face revealed that 16.8% of facial traits were significantly different between patients with BD and healthy controls. Remarkably, the patterns of facial differences were sex-specific, suggesting that BD has a different effect on male and female patients. These findings show that local facial differences could be used as biomarkers for an improved diagnosis of BD and raise awareness on the importance of studying sex differences on neurodevelopmental disorders to develop more specific and efficient treatments

High Potential of Facial Biomarkers to Diagnose Psychotic Disorders

Schizophrenia (SCZ) and Bipolar Disorder (BP) are severe psychiatric disorders (PD) that affect more than 3% of the world's population and are among the leading causes of disability worldwide. Current diagnostic systems represent these PD as independent categorical entities. However, recent studies propose that both disorders would be two different manifestations of the same psychotic spectrum continuum. Differential diagnosis is mainly based on their clinical presentation, and reliable biomarkers remain an unmet clinical need. Since the brain and the face are derived from the same ectodermal origins and their development is intimately integrated through common genetic signaling, facial biomarkers emerge as one of the most promising biological risk factors for PD. Here, we assessed the potential of facial anatomy in predicting the diagnosis of SCZ and BP. Analyses were performed in a sample of 180 adults distributed in three groups of BP patients (n=46), SCZ patients (n=67), and CNT (n=67) matched by age and premorbid IQ. Faces were manually annotated from reconstructions of magnetic resonance scans. Facial shape correctly discriminated patients with BP and SCZ, even when facial differences between patients and CNT were so subtle that are not recognizable to the untrained eye or by exploratory multivariate statistical techniques. After cross-validation, 62-65% of patients were correctly diagnosed based on face shape. This percentage is similar to the discriminatory power of other genetic and brain biomarkers. Using Artificial Neural Networks, we tested a machine learning algorithm based on facial morphology to diagnose SCZ. The overall accuracy in diagnostic classification was greater than 90%, whereas the precision ranged between 70-95% depending on the model. We also trained a Support Vector Machine classification algorithm to diagnose BP. Results showed that BP is harder to diagnose from facial biomarkers than SCZ, achieving a 72% accuracy. Euclidean Distance Matrix Analysis (EDMA) detected local facial differences involving the eyes, nose and mouth, and the relative separation/position between them. Facial anomalies were more abundant in SCZ patients, with 43-48% distances across the whole face significantly different from control subjects. In BP, the percentage of facial anomalies was lower, 24-32%, especially in women. Some facial differences were common to SCZ and BP, although the sense of change could be different among disorders. Remarkably, EDMA showed facial patterns that are disorder and gender-specific. These results demonstrate that an analysis of the spectrum of psychotic disorders under a gender perspective is crucial to further understand these disorders and to identify reliable biomarkers that can lead to early PD diagnosis

Evolutionary patterns of the human skull. A quantitative genetic analysis of craniofacial phenotypic variation

[eng] This thesis is the final outcome of the project "Quantitative genetics of craniofacial traits: a functional approach to heritability", which received support from the Wenner-Gren Foundation for Anthropological Research in 2004. The main goal is to integrate geometric morphometric with quantitative genetics in order to estimate the genetic variation underlying skull morphology and to assess its capability to evolve. The analyses herein are based on a sample of human skulls from Hallstatt, an Austrian village from the Alps. The uniqueness of this sample for evolutionary anthropological studies is the availability of associated genealogical data. The results show that substantial amounts of genetic variation underlying both size and shape and pervasive genetic integration are the two main aspects that characterize the genetic architecture of the human skull. The main developmental regions of the human skull (namely the face, the neurocranium and the basicranium) have similar amounts of genetic variation. There is evidence for genetic constraints, which reduce the evolutionary potential of the human skull. These correspond to shape features that can not evolve because they do not have sufficient genetic variation. The ability to evolve is restricted by complex patterns of covariation among cranial regions which direct evolution towards certain trajectories of morphological change that would maintain an operational and functional skull shape. Simulation analyses suggest a re-interpretation of the selective scenarios for human evolution. The origin of any one of the derived characters of modern humans may have facilitated the evolution of the others. The morphological changes associated with bipedalism may have enhanced the evolution of a more globular and expanded neurocranial shape, which could be favoured afterwards by selection for bigger and more complex brains. Natural selection has significantly acted over the last 200 years, since strong directional selection on skull shape and weak stabilizing selection on skull size has been detected at Hallstatt's population. However, other microevolutionary forces contributed to the evolution of skull morphology but in opposite directions, causing a non correspondence between secular trends and the response to selection patterns. The skull responds to these pressures through complex and widespread networks of genetic and epigenetic interactions.[cat] Aquesta tesi és el resultat final d'un projecte titulat "Quantitative genetics of craniofacial traits: a functional approach to heritability", que va rebre finançament per part de la Wenner Gren Foundation for Anthropological Research l'any 2004. El principal objectiu d'aquest projecte és integrar els mètodes de Morfometria Geomètrica i de Genètica Quantitativa per quantificar la variació genètica que determina la morfologia del crani humà i estimar la seva capacitat d'evolucionar. Les anàlisis realitzades estan basades en una mostra de cranis moderns de Hallstatt, una localitat dels Alps austríacs. Aquesta és una mostra única per a estudis d'antropologia evolutiva perquè els cranis tenen informació demogràfica i genealògica associada. Altres objectius específics de la tesi es detallen a continuació: 1) Quantificar els patrons de variació-covariació genètica, fenotípica i ambiental de la morfologia craniofacial humana, a través de caràcters craneomètrics univariats i multivariats. 2) Analitzar els patrons d'integració morfològica del crani humà, tant a nivell fenotípic com genètic. 3) Estimar la capacitat evolutiva del crani humà. 4) Simular l'evolució dels caràcters derivats de la morfologia craniofacial dels humans moderns. 5) Detectar l'acció de la selecció natural en el crani humà, combinant dades demogràfiques d'èxit reproductiu amb dades morfològiques. Els resultats obtinguts evidencien que els dos aspectes que caracteritzen l'arquitectura genètica del crani humà són, d'una banda, els elevats nivells de variació genètica que determinen tant la forma com la grandària del crani humà; i per l'altra, els patrons dominants d'integració morfològica. Les tres regions principals del crani (la cara, el neurocrani i el basicrani) presenten nivells similars de variació genètica, però la base del crani és la que mostra una major integració. Les anàlisis de Genètica Quantitativa indiquen l'existència de límits genètics al canvi morfològic, que redueixen la capacitat de resposta a la selecció. Aquests límits corresponen a característiques morfològiques que no poden evolucionar perquè no tenen suficient variació genètica heretable. La capacitat evolutiva del crani humana està restringida i dirigida cap a determinades trajectòries de canvi morfològic que mantindrien una forma cranial operativa i funcional. Les anàlisis de simulació de la selecció mostren que l'origen de qualsevol dels caràcters derivats dels humans moderns pot haver facilitat l'evolució dels altres, fet que suggereix una reinterpretació dels escenaris selectius de l'evolució humana. Concretament, els resultats indiquen que l'evolució del bipedisme podria haver estimulat l'evolució d'una volta cranial més gran i més globular, que posteriorment podria haver estat afavorida per la selecció per un cervell de major grandària i més complex, tal com indiquen les evidències moleculars. Finalment, s'ha detectat que la selecció natural ha operat en l'evolució de la forma del crani de la població de Hallstatt durant els últims 200 anys. Els resultats mostren una acció significativa de selecció direccional en la forma del crani i de selecció estabilitzadora en la grandària del crani. No obstant això, es detecta que altres forces microevolutives (flux gènic, mestissatge, variació ambiental) han participat en aquest procés evolutiu, però en direccions oposades a les seleccionades. La conclusió general d'aquesta tesi posa de manifest que el crani humà es troba sota l'acció de nombroses forces evolutives, que actuen simultàniament i dirigint el canvi morfològic. El crani respon a aquestes pressions a través de complexes xarxes d'interacció genètica i epigenètica

Quantification of Gene expression patterns to reveal the origins of abnormal morphogenesis

The earliest developmental origins of dysmorphologies are poorly understood in many congenital diseases. They often remain elusive because the first signs of genetic misregulation may initiate as subtle changes in gene expression, which are hard to detect and can be obscured later in development by secondary effects. Here, we develop a method to trace back the origins of phenotypic abnormalities by accurately quantifying the 3D spatial distribution of gene expression domains in developing organs. By applying Geometric Morphometrics to 3D gene expression data obtained by Optical Projection Tomography, we determined that our approach is sensitive enough to find regulatory abnormalities that have never been detected previously. We identified subtle but significant differences in the gene expression of a downstream target of a Fgfr2 mutation associated with Apert syndrome, demonstrating that these mouse models can further our understanding of limb defects in the human condition. Our method can be applied to different organ systems and models to investigate the etiology of malformations

Altered Genetic Expression Disrupts Facial Ontogenetic Trajectory in Down Syndrome and DYRK1A Haploinsufficiency

The development of the human face is a complex, dynamic and coordinated process. Disruptions to this ontogenetic process through genetic and/or environmental factors can lead to altered patterns of growth and development, causing facial dysmorphogenesis. The Dual-specificity tyrosine-(Y)-phosphorylation-regulated kinase 1 A (DYRK1A) is a gene located in HSA21, which over- or under-expression is associated with cognitive impairment, facial and bone dysmorphologies, as those of Down syndrome (DS) and DYRK1A haploinsufficiency syndrome (DYRK1A). In this study, we assessed facial postnatal development and morphological variation using Geometric Morphometric methods, in a cross-sectional sample of 373 children between 0 and 18 years old, including individuals diagnosed with DS (n=62) and DYRK1A (n=14), as well as control individuals (n=297). Three-dimensional facial images were obtained using a multi-camera system and further characterization of facial shape was performed recording the 3D coordinates of 21 anatomical landmarks. Results showed a clear separation of DS, DYRK1A and control groups based on their facial shape across all stages, showing a phenotypic continuum from reduced to increased levels of DYRK1A expression. Facial differences were already present at infancy (0-5 years old) and involved changes in facial retraction, mandibular protrusion, as well as changes in the relative position of the mouth, nose and eyes. Sexual dimorphism was a significant factor after adolescence within the control group (P-value<0.001), as well as within the DS (P-value=0.04) and DYRK1A groups (P-value=0.03). Euclidean Distance Matrix Analysis (EDMA) showed that 74.7% of facial traits were significantly different in individuals with DS. These localized facial traits causing differences in DS changed over postnatal development until the adult facial phenotype was achieved. Finally, multivariate regression analysis of facial shape on age (% predicted=13.26%; P-value<0.0001) showed differences in the intercept and slope of the ontogenetic slopes of the three groups. These results highlight the processes by which these genetic syndromes significantly alter normal facial postnatal growth, increasing facial differences already established before birth. Our study suggests that changes in heterochrony can drive modifications in the rate or timing of facial development and produce morphological variation

Multimodal in vivo imaging of the integrated postnatal development of brain and skull and its co-modulation with neurodevelopment in a Down syndrome mouse model

The brain and skeletal systems are intimately integrated during development through common molecular pathways. This is evidenced by genetic disorders where brain and skull dysmorphologies are associated. However, the mechanisms underlying neural and skeletal interactions are poorly understood. Using the Ts65Dn mouse model of Down syndrome (DS) as a case example, we performed the first longitudinal assessment of brain, skull and neurobehavioral development to determine alterations in the coordinated morphogenesis of brain and skull. We optimized a multimodal protocol combining in vivo micro-computed tomography (μCT) and magnetic resonance imaging (μMRI) with morphometric analyses and neurodevelopmental tests to longitudinally monitor the different systems' development trajectories during the first postnatal weeks. We also explored the impact of a perinatal treatment with green tea extracts enriched in epigallocatechin-3-gallate (GTE-EGCG), which can modulate cognition, brain and craniofacial development in DS. Our analyses quantified alterations associated with DS, with skull dysmorphologies appearing before brain anomalies, reduced integration and delayed acquisition of neurodevelopmental traits. Perinatal GTE-EGCG induced disparate effects and disrupted the magnitude of integration and covariation patterns between brain and skull. Our results exemplify how a longitudinal research approach evaluating the development of multiple systems can reveal the effect of morphological integration modulating the response of pathological phenotypes to treatment, furthering our understanding of complex genetic disorders